Fibronectin (FN) was first reported by Morrison et al. [Morrison, P. R. et al., J. Am. Chem. Soc., 70, 3103 (1948)] as one of plasma proteins in 1948. Being a multifunctional protein broadly distributed in various tissues and body fluids, this substance is known to be involved, as a cell adhesion factor, in a large variety of biological events such as the migration, differentiation, proliferation and canceration of cells [Sekiguchi, K.: Cell Engineering, 4 (6), 485-497 (1985)].
Meanwhile, FN as it is synthesized in the liver and occurring in the blood is known as plasma FN (pFN), and FN as it is detected on the tissue and cell surfaces is called cellular FN (cFN) [Sekiguchi et al., J. Biol. Chem., 260 (8) 5105-5114 (1985)]. It has been shown that these species of FN are subject to molecular diversity due to alternative splicing of the early gene transcription product. As the regions subject to such alternative splicing, there are three regions called EDA, EDB and IIIcs, and it is believed that a large number of molecular species occur according to varied combinations of expression of these regions. In pFN, the above-mentioned EDA and EDB regions have not been appreciably expressed. On the other hand, cFN is an FN with a high degree of expression of said EDA region. Peters J. H. et al. conjugated a peptide having 29 amino acids with keyhole limpet hemocyanin (KLH) to prepare an immunogen, constructed an anti-cFN polyclonal antibody specifically reactive to cFN having the EDA region, and using the antibody, demonstrated that the normal human blood contains traces of cFN and that the blood cFN level is markedly elevated in patients with collagen disease accompanied by vasculitis [Am. Rev. Respir. Dis., 138, 167-174 (1988); J. Lab. Clin. Med., 113 (5), 586-597 (1989)]. According to the above report of Peters et al., the physiological implications of increased blood cFN in patients with collagen disease accompanied by vasculitis are not definitely known but a correlation between the stage or severity of disease and the blood concentration of cFN is suggested.
The investigation made by the inventors of the present invention revealed that whereas the plasma concentration of cFN in healthy humans is 1 to 2 .mu.g/ml, it is as high as 10 and odd .mu.g/ml in patients with various diseases. On the other hand, pFN is invariably within the normal range of 270 to 400 .mu.g/ml, irrespective of subjects.
Therefore, it is a subject of intriguing interest to investigate how the course of disease is modified by selective elimination of cFN from the blood of patients with collagen disease presenting with abnormal elevations of cFN. The recent study by the inventors of the present invention uncovered that when the plasma of a patient with rheumatoid arthritis is subjected to cryofiltration, cFN is partially removed from the plasma, resulting in relief of the symptoms. However, the cryofiltration procedure is disadvantageous in that it involves discarding of the useful plasma protein in large quantities and is not capable of selective elimination of cFN.
The inventors of the present invention previously developed a method for separating and eliminating cFN selectively from the patient's blood and a selective cFN adsorbent for the elimination thereof and filed a patent application (PCT/JP92/00359). It was, however, subsequently discovered that, occurring as granules or gels, the cFN adsorbent disclosed in the above application has the drawback that it incurs a high production cost, must be maintained in wet condition and, in packing it into an adsorption module column for extra-corporeal circulation, must be handled as floated on water.
In view of the above disadvantages, the inventors conducted further intensive research and established a high-sensitivity protocol for assay of cFN using a previously developed monoclonal antibody which recognizes the amino acid sequence of the EDA region of FN. The inventors further discovered that a nonwoven fabric made of cellulose sulfate fiber has a selective affinity for cFN. Thus, it was discovered that such a nonwoven cellulose sulfate fabric adsorbs cFN and, because of its difference in affinity between cFN and pFN, adsorbs cFN with greater specificity. Based on these findings, the inventors further explored and developed a new technology which enabled fractionation between pFN and cFN. The present invention has been developed on the basis of the above findings.